The present invention relates to pyrrolo[2,3-d]pyrimidine compounds which are inhibitors of protein tyrosine kinases, such as the enzyme Janus Kinase 3 (hereinafter also referred to as JAK3) and as such are useful therapy as immunosuppressive agents for organ transplants, lupus, multiple sclerosis, rheumatoid arthritis, psoriasis, Type I diabetes and complications from diabetes, cancer, asthma, atopic dermatitis, autoimmune thyroid disorders, ulcerative colitis, Crohn""s disease, Alzheimer""s disease, Leukemia and other indications where immunosuppression would be desirable.
This invention also relates to a method of using such compounds in the treatment of the above indications in mammals, especially humans, and the pharmaceutical compositions useful therefor.
JAK3 is a member of the Janus family of protein tyrosine kinases. Although the other members of this family are expressed by essentially all tissues, JAK3 expression is limited to hematopoetic cells. This is consistent with its essential role in signaling through the receptors for IL-2, IL-4, IL-7, IL-9 and IL-15 by non-covalent association of JAK3 with the gamma chain common to these multichain receptors. XSCID patient populations have been identified with severely reduced levels of JAK3 protein or with genetic defects to the common gamma chain, suggesting that immunosuppression should result from blocking signaling through the JAK3 pathway. Animal studies have suggested that JAK3 not only plays a critical role in B and T lymphocyte maturation, but that JAK3 is constitutively required to maintain T cell function. Modulation of immune activity through this novel mechanism can prove useful in the treatment of T cell proliferative disorders such as transplant rejection and autoimmune diseases.
The present invention relates to a compound of the formula 
or the pharmaceutically acceptable salt thereof; wherein
R1 is a group of the formula
or when n is at least 1, D and E, or D and X, are each CR7R8, the adjacent R7 groups may be taken together, with the carbons to which they are attached, to form groups of the formulas 
xe2x80x83wherein the dashed lines represent optional double bonds;
a is 0, 1 or 2;
m, A, B and X are as defined above; and
G, J, L and M are each independently oxygen, S(O)d wherein d is 0, 1 or 2, NR6 or CR7R8 wherein R6, R7 and R8 are as defined above;
or when n is 1, D and E are each CR7R8 and m is 1, A and B are each CR7R8, the respective adjacent R7 groups may be taken together, with the carbons to which they are attached, to form a group of the formula 
wherein the dashed bond represent optional double bonds;
a, G, J. L and M are as define above;
r is 0 or 1;
c is 0, 1 or 2; and
R, W, Y and S are each independently oxygen, S(O)d wherein d is 0, 1 or 2, NR6 or CR7R8 wherein R6, R7 and R8 are as defined above;
R2 and R3 are each independently selected from the group consisting of hydrogen, deuterium, amino, halo, hydoxy, nitro, carboxy, (C2-C6)alkenyl, (C2-C6)alkynyl, trifluoromethyl, 
wherein the dashed line represents optional double bonds;
m is 0, 1, 2 or 3;
n is 0, 1, 2 or 3;
X, B and D are each independently oxygen, S(O)d wherein d is 0, 1 or 2, NR6 or CR7R8;
A and E are each CR7R8; and
R6 is selected from the group consisting of hydrogen, (C1-C6)alkyl, trifluoromethyl, trifluoromethyl(C1-C6)alkyl, (C1-C6)alkyl(difluoromethylene), (C1-C3)alkyl (difluoromethylene)(C1-C3)alkyl, (C1-C6)alkoxy(C1-C6)acyl, (C1-C6)alkylamino(C1-C6)acyl, ((C1-C6)alkyl)2amino(C1-C6)acyl, (C6-C10)aryl, (C5-C9)heteroaryl, (C6-C10)aryl(C1-C6)alkyl, (C5-C9)heteroaryl(C1-C6)acyl, (C6-C10)aryl(C6-C10)aryl, (C6-C10)aryl(C6-C10)aryl(C1-C6)alkyl, (C3-C6)cycloalkyl, (C3-C6)cycloalkyl(C1-C6)alkyl, hydroxy(C2-C6)alkyl, (C1-C6)acyloxy(C2-C6)alkyl, (C1-C6)alkoxy(C2-C6)alkyl, piperazinyl(C1-C6)alkyl, (C1-C6)acylamino(C1-C6)alkyl, (C6-C10)aryl(C1-C6)alkoxy(C1-C6)alkyl, (C5-C9)heteroaryl(C1-C6)alkoxy(C1-C6)alkyl, (C1-C6)alkylthio(C1-C6)alkyl, (C6-C10)arylthio(C1-C6)alkyl, (C1-C6)alkylsulfinyl(C1-C6)alkyl, (C6-C10)arylsulfinyl(C1-C6)alkyl, (C1-C6)alkylsulfonyl(C1-C6)alkyl, (C6-C10)arylsulfonyl(C1-C6)alkyl, amino(C1-C6)alkyl, (C1-C6)alkylamino(C1-C6)alkyl, ((C1-C6)alkyl)2amino(C1-C6)alkyl, R13CO(C1-C6)alkyl wherein R13 is R20O or R20R21N wherein R20 and R21 are each independently selected from the group consisting of hydrogen, (C1-C6)alkyl, (C6-C10)aryl(C1-C6)alkyl or (C5-C9)heteroaryl(C1-C6)alkyl; or R14(C2-C6)alkyl wherein R14 is (C1-C6)acylpiperazino, (C6-C10)arylpiperazino, (C5-C9)heteroarylpiperazino, (C1-C6)alkylpiperazino, (C6-C10)aryl(C1-C6)alkylpiperazino, (C5-C9)heteroaryl(C1-C6)alkylpiperazino, morpholino, thiomorpholino, piperidino, pyrrolidino, piperidyl, (C1-C6)alkylpiperidyl, (C6-C10)arylpiperidyl, (C5-C9)heteroarylpiperidyl, (C6-C10)aryl(C1-C6)alkylpiperidyl, (C5-C9)heteroaryl(C1-C6)alkylpiperidyl, (C1-C6)alkoxyacyl, (C1-C6)alkylaminoaryl, ((C1-C6)alkyl2aminoacyl or (C1-C6)acylpiperidyl;
R7 and R8 are each independently selected from the group consisting of hydrogen, deuterium, (C1-C6)alkyl, amino, hydroxy, (C1-C6)alkoxy, (C1-C6)alkylamino, ((C1-C6)alkyl)amino, (C1-C6)acylamino, (C1-C6)acyl(C1-C6)alkylamino, carboxy, (C1-C6)alkoxyacyl, (C1-C6)alkylaminoacyl, ((C1-C6)alkyl)2aminoacyl, aminoacyl, trifluoromethyl, trifluoromethyl(C1-C6)alkyl, (C1-C6)alkyl (difluoromethylene), (C1-C3)alkyl(difluoromethylene)(C1-C3)alkyl, (C6-C10)aryl, (C5-C9)heteroaryl, (C6-C10)aryl(C1-C6)alkyl, (C5-C9)heteroaryl(C1-C6)alkyl, (C6-C10)aryl(C6-C10)aryl, (C6-C10)aryl(C6-C10)aryl(C1-C6)alkyl, (C3-C6)cycloalkyl, (C3-C6)cycloalkyl(C1-C6)alkyl, hydroxy(C1-C6)alkyl, (C1-C6)acyloxy(C1-C6)alkyl, (C1-C6)alkoxy(C1-C6)alkyl, piperazinyl(C1-C6)alkyl, (C1-C6)acylamino(C1-C6)alkyl, piperidyl, (C1-C6)alkylpiperidyl, (C6-C10)aryl(C1-C6)alkoxy(C1-C6)alkyl, (C5-C9)heteroaryl (C1-C6)alkoxy(C1-C6)alkyl, (C1-C6)alkylthio(C1-C6)alkyl, (C6-C10)arylthio(C1-C6)alkyl, (C1-C6)alkylsulfinyl(C1-C6)alkyl, (C6-C10)arylsulfinyl(C1-C6)alkyl, (C1-C6)alkylsulfonyl(C1-C6)alkyl, (C6-C10)arylsulfonyl(C1-C6)alkyl, amino(C1-C6)alkyl, (C1-C6)alkylamino(C1-C6)alkyl, ((C1-C6)alkyl)2amino(C1-C6)alkyl, R13CO(C1-C6)alkyl or R13CO(C3-C10)cycloalkyl wherein R13 is R20O or R20R21N wherein R20 and R21 are each independently selected from the group consisting of hydrogen, (C1-C6)alkyl, (C6-C10)aryl(C1-C6)alkyl or (C5-C9)heteroaryl(C1-C6)alkyl; R14, R14(C1-C6)alkyl or R14(C3-C10)cycloalkyl wherein R14 is (C1-C6)acylpiperazino, (C6-C10)arylpiperazino, (C5-C9)heteroarylpiperazino, (C1-C6)alkylpiperazino, (C6-C10)aryl(C1-C6)alkylpiperazino, (C5-C9)heteroaryl(C1-C6)alkylpiperazino, morpholino, thiomorpholino, piperidino, pyrrolidino, piperidyl, (C1-C6)alkylpiperidyl, (C6-C10)arylpiperidyl, (C5-C9)heteroarylpiperidyl, (C6-C10)aryl(C1-C6)alkylpiperidyl, (C5-C9)heteroaryl(C1-C6)alkylpiperidyl or (C1-C6)acylpiperidyl; or a group of the formula 
wherein p is 0, 1, 2 or 3; and
Z is hydroxy, (C1-C6)alkoxy or NR1R2 wherein R1 and R2 are each independently selected from the group consisting of hydrogen, (C1-C6)alkyl, piperidyl, (C1-C6)alkylpiperidyl, (C6-C10)arylpiperidyl, (C5-C9)heteroarylpiperidyl, (C6-C10)aryl(C1-C6)alkylpiperidyl, (C5-C9)heteroaryl(C1-C6)alkylpiperidyl, (C1-C6)acylpiperidyl, (C6-C10)aryl, (C5-C9)heteroaryl, (C6-C10)aryl(C1-C6)alkyl, (C5-C9)heteroaryl(C1-C6)alkyl, (C6-C10)aryl(C6-C10)aryl, (C6-C10)aryl(C6-C10)aryl(C1-C6)alkyl, (C3-C6)cycloalkyl, (C3-C6)cycloalkyl(C1-C6)alkyl, R5(C1-C6)alkyl, (C1-C5)alkyl(CHR5)(C1-C6)alkyl wherein R5 is hydroxy, (C1-C6)acyloxy, (C1-C6)alkoxy, piperazino, (C1-C6)acylamino, (C1-C6)alkylthio, (C6-C10)arylthio, (C1-C6)alkylsulfinyl, (C6-C10)arylsulfinyl, (C1-C6)alkylsulfoxyl, (C6-C10)arylsulfoxyl, amino, (C1-C6)alkylamino, ((C1-C6)alkyl)2 amino, (C1-C6)acylpiperazino, (C1-C6)alkylpiperazino, (C6-C10)aryl(C1-C6)alkylpiperazino, (C5-C9)heteroaryl(C1-C6)alkylpiperazino, morpholino, thiomorpholino, piperidino or pyrrolidino; R6(C1-C6)alkyl, (C1-C5)alkyl(CHR6)(C1-C6)alkyl wherein R6 is piperidyl, (C1-C6)alkylpiperidyl, (C6-C10)arylpiperidyl, (C6-C10)aryl(C1-C6)alkylpiperidyl, (C5-C9)heteroarylpiperidyl or (C5-C9)heteroaryl(C1-C6)alkylpiperidyl;
trifluoromethoxy, (C1-C6)alkyl, (C1-C6)alkoxy wherein the alkyl or alkoxy groups are optionally substittued by one to three groups selected from halo, hydroxy, carboxy, amino (C1-C6)alkylthio, (C1-C6)alkylamino, ((C1-C6)alkyl)2amino, (C5-C9)heteroaryl, (C2-C9)heterocycloalkyl, (C3-C9)cycloalkyl or (C6-C10)aryl; or R2 and R3 are each independently (C3-C10)cycloalkyl, (C3-C10)cycloalkoxy, (C1-C6)alkylamino, ((C1-C6)alkyl)2amino, (C6-C10)arylamino, (C1-C6)alkylthio, (C6-C10)arylthio, (C1-C6)alkylsulfinyl, (C6-C10)arylsulfinyl, (C1-C6)alkylsulfonyl, (C6-C10)arylsulfonyl, (C1-C6)acyl, (C1-C6)alkoxy-COxe2x80x94NHxe2x80x94, (C1-C6)alkyamino-COxe2x80x94, (C5-C9)heteroaryl, (C2-C9)heterocycloalkyl or (C6-C10)aryl wherein the heteroaryl, heterocycloalkyl and aryl groups are optionally substituted by one to three halo, (C1-C6)alkyl, (C1-C6)alkyl-COxe2x80x94NHxe2x80x94, (C1-C6)alkoxy-COxe2x80x94NHxe2x80x94, (C1-C6)alkyl-COxe2x80x94NHxe2x80x94(C1-C6)alkyl, (C1-C6)alkoxy-COxe2x80x94NHxe2x80x94(C1-C6)alkyl, (C1-C6)alkoxy-COxe2x80x94NHxe2x80x94(C1-C6)alkoxy, carboxy, carboxy(C1-C6)alkyl, carboxy(C1-C6)alkoxy, benzyloxycarbonyl(C1-C6)alkoxy, (C1-C6)alkoxycarbonyl(C1-C6)alkoxy, (C6-C10)aryl, amino, amino(C1-C6)alkyl, (C1-C6)alkoxycarbonylamino, (C6-C10)aryl(C1-C6)alkoxycarbonylamino, (C1-C6)alkylamino, ((C1-C6)alkyl)2amino, (C1-C6)alkylamino(C1-C6)alkyl, ((C1-C6)alkyl)2amino(C1-C6)alkyl, hydroxy, (C1-C6)alkoxy, carboxy, carboxy(C1-C6)alkyl, (C1-C6)alkoxycarbonyl, (C1-C6)alkoxycarbonyl(C1-C6)alkyl, (C1-C6)alkoxy-COxe2x80x94NHxe2x80x94, (C1-C6)alkyl-COxe2x80x94NHxe2x80x94, cyano, (C5-C9)heterocycloalkyl, amino-COxe2x80x94NHxe2x80x94, (C1-C6)alkylamino-COxe2x80x94NHxe2x80x94, ((C1-C6)alkyl)2amino-COxe2x80x94NHxe2x80x94, (C6-C10)arylamino-COxe2x80x94NHxe2x80x94, (C5-C9)heteroarylamino-COxe2x80x94NHxe2x80x94, (C1-C6)alkylamino-COxe2x80x94NHxe2x80x94(C1-C6)alkyl, ((C1-C6)alkyl)2amino-COxe2x80x94NHxe2x80x94(C1-C6)alkyl, (C6-C10)arylamino-COxe2x80x94NHxe2x80x94(C1-C6)alkyl, (C5-C9)heteroarylamino-COxe2x80x94NHxe2x80x94(C1-C6)alkyl, (C1-C6)alkylsulfonyl, (C1-C6)alkylsulfonylamino, (C1-C6)alkylsulfonylamino(C1-C6)alkyl, (C6-C10)arylsulfonyl, (C6-C10)arylsulfonylamino, (C6-C10)arylsulfonylamino(C1-C6)alkyl, (C1-C6)alkylsulfonylamino, (C1-C6)alkylsulfonylaminc(C1-C6)alkyl, (C5-C9)heteroaryl or (C2-C9)heterocycloalkyl;
with the proviso that when A, B or X, in formulas V or VI, is defined as NR6 or CR7R8, R2 and/or R3 must be halo;
with the proviso that when R2 and R3 are each independently hydrogen or (C1-C6)alkyl, R1 cannot be unsubstituted piperidinyl;
with the proviso that when R2 and R3 are each hydrogen, R1 cannot be unsubstituted mopholinyl or pyrrolidinyl;
with the proviso that when R2 and R3 are each hydrogen, R1 cannot be piperazinyl; and
with the proviso that the groups of formulas IV, V, VI or XIII do not contain two or more oxygens, sulfurs or combinations thereof in adjacent positions.
The present invention also relates to the pharmaceutically acceptable acid addition salts of compounds of the formula I. The acids which are used to prepare the pharmaceutically acceptable acid addition salts of the aforementioned base compounds of this invention are those which form non-toxic acid addition salts, i.e., salts containing pharmacologically acceptable anions, such as the hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfate, phosphate, acid phosphate, acetate, lactate, citrate, acid citrate, tartrate, bitartrate, succinate, maleate, fumarate, gluconate, saccharate, benzoate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate and pamoate [i.e., 1,1xe2x80x2-methylene-bis-(2-hydroxy-3-naphthoate)]salts.
The invention also relates to base addition salts of formula 1. The chemical bases that may be used as reagents to prepare pharmaceutically acceptable base salts of those compounds of formula I that are acidic in nature are those that form non-toxic base salts with such compounds. Such non-toxic base salts include, but are not limited to those derived from such pharmacologically acceptable cations such as alkali metal cations (eg, potassium and sodium) and alkaline earth metal cations (e.g., calcium and magnesium), ammonium or water-soluble amine addition salts such as N-methylglucamine-(meglumine), and the lower alkanolammonium and other base salts of pharmaceutically acceptable organic amines.
The term xe2x80x9calkylxe2x80x9d, as used herein, unless otherwise indicated, includes saturated monovalent hydrocarbon radicals having straight, branched or cyclic moieties or combinations thereof.
The term xe2x80x9calkoxyxe2x80x9d, as used herein, includes 0-alkyl groups wherein xe2x80x9calkylxe2x80x9d is defined above.
The term xe2x80x9chaloxe2x80x9d, as used herein, unless otherwise indicated, includes fluoro, chloro, bromo or iodo.
The compounds of this invention may contain double bonds. When such bonds are present, the compounds of the invention exist as cis and trans configurations and as mixtures thereof.
Unless otherwise indicated, the alkyl and alkenyl groups referred to herein, as well as the alkyl moieties of other groups referred to herein (eg, alkoxy), may be linear or branched, and they may also be cyclic (e.g., cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl or cycloheptyl) or be linear or branched and contain cyclic moieties. Unless otherwise indicated, halogen includes fluorine, chlorine, bromine, and iodine.
(C3-C10)Cycloalkyl when used herein refers to cycloalkyl groups containing zero to two levels of unsaturation such as cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, 1,3-cyclohexadiene, cycloheptyl, cycloheptenyl, bicyclo[3.2.1]octane, norbornanyl etc.
(C2-C9)Heterocycloalkyl when used herein refers to pyrrolidinyl, tetrahydrofuranyl, dihydrofuranyl tetrahydropyranyl, pyranyl, thiopyranyl, aziridinyl, oxiranyl, methylenedioxyl, chromenyl, isoxazolidinyl, 1,3-oxazolidin-3-yl, isothiazolidinyl, 1,3-thiazolidin-3-yl, 1,2-pyrazolidin-2-yl, 1,3-pyrazolidin-1-yl, piperidinyl, thiomorpholinyl, 1,2-tetrahydrothiazin-2-yl, 1,3-tetrahydrothiazin-3-yl, tetrahydrothiadiazinyl, morpholinyl, 1,2-tetrahydrodiazin-2-yl, 1,3-tetrahydrodiazin-3-yl, tetrahydroazepinyl, piperazinyl, chromanyl, etc. One of ordinary skill in the art will understand that the connection of said (C2-C9)heterocycloalkyl rings is through a carbon or a sp3 hybridized nitrogen heteroatom.
(C2-C9)Heteroaryl when used herein refers to furyl, thienyl, thiazolyl, pyrazolyl, isothiazolyl, oxazolyl, isoxazolyl, pyrrolyl, triazolyl, tetrazolyl, imidazolyl, 1,3,5-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,3-oxadiazolyl, 1,3,5-thiadiazolyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, pyridyl, pyrimidyl, pyrazinyl, pyridazinyl, 1,2,4-triazinyl, 1,2,3-triazinyl, 1,3,5-triazinyl, pyrazolo[3,4-b]pyridinyl, cinnolinyl, pteridinyl, purinyl, 6,7-dihydro-5H-[1]pyrindinyl, benzo[b]thiophenyl, 5,6,7,8-tetrahydro-quinolin-3-yl, benzoxazolyl, benzothiazolyl, benzisothiazolyl, benzisoxazolyl, benzimidazolyl, thianaphthenyl, isothianaphthenyl, benzofuranyl, isobenzofuranyl, isoindolyl, indolyl, indolizinyl, indazolyl, isoquinolyl, quinolyl, phthalazinyl, quinoxalinyl, quinazolinyl, benzoxazinyl; etc. One of ordinary skill in the art will understand that the connection of said (C2-C9)heteroaryl rings is through a carbon atom or a sp3 hybridized nitrogen heteroatom.
(C6-C10)aryl when used herein refers to phenyl or naphthyl.
Compounds of formula (I) may be administered in a pharmaceutically acceptable form either alone or in combination with one or more additional agents which modulate a mammalian immune system or with antiinflammatory agents. These agents may include but are not limited to cyclosporin A (e.g. Sandimmune(copyright) or Neoral(copyright), rapamycin, FK-506 (tacrolimus), leflunomide, deoxyspergualin, mycophenolate (e.g. Cellcept(copyright)), azathioprine (e.g. Imuran(copyright)), daclizumab (e.g. Zenapax(copyright), OKT3 (e.g. Orthoclone(copyright)), AtGam, aspirin, acetaminophen, ibuprofen, naproxen, piroxicam, and antiinflammatory steroids (e.g. prednisolone or dexamethasone). These agents may be administered as part of the same or separate dosage forms, via the same or different routes of administration, and on the same or different administration schedules according to standard pharmaceutical practice.
The compounds of this invention include all configurational isomers (es, cis and trans isomers) and all optical isomers of compounds of the formula I (e., enantiomers and diastereomers), as well as racemic, diastereomeric and other mixtures of such isomers. This invention also includes all rotamers of compounds of formula I as well as scelemic mixtures.
Preferred compounds of formula I include those wherein R1 is is a group of the formula 
wherein the dashed line represents optional double bonds;
m is 0, 1, 2 or 3;
n is 0, 1, 2 or 3;
X, B and D are each independently oxygen, S(O)d wherein d is 0, 1 or 2, NR6 or CR7R8;
A and E are each independently CR7R8or NR6;
or when n is 1, D and E are each CR7R8 and m is 1, A and B are each CR7R8, the respective adjacent R7 groups may be taken together, with the carbons to which they are attached, to form a group of the formula 
wherein the dashed bond represent optional double bonds;
a, G, J. L and M are as define above;
r is 0 or 1;
c is 0, 1 or 2; and
R, W, Y and S are each independently oxygen, S(O)d wherein d is 0, 1 or 2, NR6 or CR7R8 wherein R6, R7 and R8 are as defined above.
Other preferred compounds of formula I include those wherein R2 and R3 are each independently selected from the group consisting of hydrogen, (C1-C6)alkyl, (C1-C6)alkoxy, (C3-C10)cycloalkyl, (C3-C10)cycloalkoxy, (C2-C9)heterocycloalkyl, (C5-C9)heteroaryl or (C6-C10)aryl.
Specific preferred compounds of formula I include the following:
5-Fluoro-4-piperidin-1-yl-7H-pyrrolo[2,3-d]pyrimidine;
4-Piperidin-1-yl-5-trifluoromethyl-7H-pyrrolo[2,3-d]pyrimidine;
2-{3-Ethyl-4-[methyl-(7H-pyrrolo(2,3-d]pyrimidin-4-yl)-aminol-cyclopentyl}-propan-2-ol;
2-{3-Ethyl-4-[(2-hydroxy-ethyl)-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-amino]-cyclopentyl}-propan-2-ol;
N,N-Dimethyl-Nxe2x80x2-[3-(4-piperidin-1-yl-7H-pyrrolo[2,3-d]pyrimidin-5-yl)-benzyl]-ethane-1,2-diamine;
2-[1-(5-m-Tolyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)-piperidin-4-yl]-ethanol;
5-(3-Isopropyl-phenyl)4-piperidin-1-yl-7H-pyrrolo[2,3-d]pyrimidine;
5-(3-Methyl-3H-imidazol-4-yl)-4-piperidin-1-yl-7H-pyrrolo[2,3-d]pyrimidine;
5-(1-Methyl-1H-imidazol-4-yl)-4-piperidin-1-yl-7H-pyrrolo[2,3-d]pyrimidine;
5-(2-Methyl-pyridin4-yl)-4-piperidin-1-yl-7H-pyrrolo[2,3-d]pyrimidine;
5-Chloro-4-piperidin-1-yl-7H-pyrrolo[2,3-d]pyrimidine;
5-Chloro-4-(octahydro-indol-1-yl)-7H-pyrrolo[2,3-d]pyrimidine;
5-Ethynyl-4-piperidin-1-yl-7H-pyrrolo[2,3-d]pyrimidine;
4-Piperidin-1-yl-5-m-tolyl-7H-pyrrolo[2,3-d]pyrimidine; and
4-(3,3-Dimethyl-piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine.
The present invention also relates to a pharmaceutical composition for (a) treating or preventing a disorder or condition selected from organ transplant rejection, lupus, multiple sclerosis, rheumatoid arthritis, psoriasis, Type I diabetes and complications from diabetes, cancer, asthma, atopic dermatitis, autoimmune thyroid disorders, ulcerative colitis, Crohn""s disease, Alzheimer""s disease, Leukemia, and other autoimmune diseases or (b) the inhibition of protein tyrosine kinases or Janus Kinase 3 (JAK3) in a mammal, including a human, comprising an amount of a compound of formula I or a pharmaceutically acceptable salt thereof, effective in such disorders or conditions and a pharmaceutically acceptable carrier.
The present invention also relates to a pharmaceutical composition for (a) treating or preventing a disorder or condition selected from organ transplant rejection, lupus, multiple sclerosis, rheumatoid arthritis, psoriasis, Type I diabetes and complications from diabetes, cancer, asthma, atopic dermatitis, autoimmune thyroid disorders, ulcerative colitis, Crohn""s disease, Alzheimer""s disease, Leukemia, and other autoimmune diseases or (b) the inhibition of protein tyrosine kinases or Janus Kinase 3 (JAK3) in a mammal, including a human, comprising an amount of a compound of formula I or a pharmaceutically acceptable salt thereof, alone or in combination with T-cell immunosuppressant or antiinflammatory agents, effective in such disorders or conditions and a pharmaceutically acceptable carrier.
The present invention also relates to a method for the inhibition of protein typrosine kinases or Janus Kinase 3 (JAK3) in a mammal, including a human, comprising administering to said mammal an effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof.
The present invention also relates to a method for treating or preventing a disorder or condition selected from organ transplant rejection, lupus, multiple sclerosis, rheumatoid arthritis, psoriasis, Type I diabetes and complications from diabetes, cancer, asthma, atopic dermatitis, autoimmune thyroid disorders, ulcerative colitis, Crohn""s disease, Alzheimer""s disease, Leukemia, and other autoimmune diseases in a mammal, including a human, comprising administering to said mammal an amount of a compound of formula I or a pharmaceutically acceptable salt thereof, effective in treating such a condition.
The present invention also relates to a method for the inhibition of protein typrosine kinases or Janus Kinase 3 (JAK3) in a mammal, including a human, comprising administering to said mammal an effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof, alone or in combination with T-cell immunosuppressant or antiinflammatory agents.
The present invention also relates to a method for treating or preventing a disorder or condition selected from organ transplant rejection, lupus, multiple sclerosis, rheumatoid arthritis, psoriasis, Type I diabetes and complications from diabetes, cancer, asthma, atopic dermatitis, autoimmune thyroid disorders, ulcerative colitis, Crohn""s disease, Alzheimer""s disease, Leukemia, and other autoimmune diseases in a mammal, including a human, comprising administering to said mammal an amount of a compound of formula I or a pharmaceutically acceptable salt thereof, alone or in combination with T-cell immunosuppressant or antiinflammatory agents, effective in treating such a condition.
The following reaction Schemes illustrate the preparation of the cormpounds of the present invention. Unless otherwise indicated R1, R2, R3 and R9 in the reaction Schemes and the discussion that follow are defined as above. 
In reaction 1 of Scheme 1, the 4-chloropyrrolo[2,3-d]pyrimidine compound of formula XVII is converted to the corresponding compound of formula XVI, wherein R is benzenesulfonyl or benzyl, by treating XVII with benzenesulfonyl chloride, benzylchloride or benzylbromide in the presence of a base, such as sodium hydride or potassium carbonate, and a polar aprotic solvent, such as dimethylformamide or tetrahydrofuran. The reaction mixture is stirred at a temperature between about 0xc2x0 C. to about 70xc2x0 C., preferably about 30xc2x0 C., for a time period between about 1 hour to about 3 hours, preferably about 2 hours.
In reaction 2 of Scheme 1, the 4-chloropyrrolo[2,3-d]pyrimidine compound of formula XVI is converted to the corresponding 4-aminopyrrolo[2,3-d]pyrimidine compound of formula XV by coupling XVI with a compound of the formula R1H. The reaction is carried out in an alcohol solvent, such as tert-butanol, methanol or ethanol, or other high boiling organic solvents, such as dimethylformamide, 1,4-dioxane or 1,2-dichloroethane, at a temperature between about 60xc2x0 C. to about 120xc2x0 C., preferably about 80xc2x0 C. Typical reaction times are between about 2 hours to about 48 hours, preferably about 16 hours.
In reaction 3 of Scheme 1, removal of the protecting group from the compound of formula XV, wherein R is benzenesulfonyl, to give the corresponding compound of formula I, is carried out by treating XV with an alkali base, such as sodium hydroxide or potassium hydroxide, in an alcohol solvent, such as methanol or ethanol, or mixed solvents, such as alcohol/tetrahydrofuran or alcohol/water. The reaction is carried out at room temperature for a time period between about 15 minutes to about 1 hour, preferably 30 minutes. Removal of the protecting group from the compound of formula XV, wherein R is benzyl, is conducted by treating XV with sodium in ammonia at a temperature of about xe2x88x9278xc2x0 C. for a time period between about 15 minutes to about 1 hour.
In reaction 1 of Scheme 2, the 4-chloropyrrolo[2,3-d]pyrimidine compound of formula XXI, wherein R is hydrogen or benzenesulfonate, is converted to the 4-chloro-5-halopyrrolo[2,3-d]pyrimidine compound of formula XX, wherein Y is chloro, bromo or iodo, by reacting XXI with N-chlorosuccinimide, N-bromosuccinimide or N-iodosuccinimide. The reaction mixture is heated to reflux, in chloroform, for a time period between about 1 hour to about 3 hours, preferably about 1 hour. Alternatively, in, reaction 1 of Scheme 2, the 4 chloropyrrolo[2,3-d]pyrimidine of formula XXI, wherein R is hydrogen, is converted to the corresponding 4-chloro-5-nitropyrrolo[2,3-d]pyrimidine of formula XX, wherein Y is nitro, by reacting XXI with nitric acid in sulfuric acid at a temperature between about xe2x88x9210xc2x0 C. to about 10xc2x0 C., preferably about 0xc2x0 C., for a time period between about 5 minutes to about 15 minutes, preferably about 10 minutes. The compound of formula XXI, wherein Y is nitro, is converted to the corresponding 4-chloro-5-aminopyrrolo[2,3-d]pyrimidine of the formula XX, wherein Y is amino, by reacting XXI under a variety of conditions known to one skilled in the art such as palladium hydrogenolysis or tin(IV)chloride and hydrochloric acid.
In reaction 2 of Scheme 2, the 4-chloro-5-halopyrrolo[2,3-d]pyrimidine compound of formula XX, wherein R is hydrogen, is converted to the corresponding compound of formula XIX, wherein R2 is (C1-C6)alkyl or benzyl, by treating XX with N-butyllithium, at a temperature of about xe2x88x9278xc2x0 C., and reacting the dianion intermediate so formed with an alkylhalide or benzylhalide at a temperature between about xe2x88x9278xc2x0 C. to room temperature, preferably room temperature. Alternatively, the dianion so formed is reacted with molecular oxygen to form the corresponding 4-chloro-5-hydroxypyrrolo[2,3-d]pyrimidine compound of formula XIX, wherein R2 is hydroxy. The compound of formula XX, wherein Y is bromine or iodine and R is benzenesulfonate, is converted to the compound of formula XIX, wherein R2 is (C6-C12)aryl or vinyl, by treating XX with N-butyllithium, at a temperature of about xe2x88x9278xc2x0 C., followed by the addition of zinc chloride, at a temperature of about xe2x88x9278xc2x0 C. The corresponding organo zinc intermediate so formed is then reacted with aryliodide or vinyl iodide in the presence of a catalytic quantity of palladium. The reaction mixture is stirred at a temperature between about 50xc2x0 C. to about 80xc2x0 C., preferably about 70xc2x0 C., for a time period between about 1 hour to about 3 hours, preferably about 1 hour.
In reaction 3 of Scheme 2, the compound of formula XIX is converted to the correspondihg compound of formula XVI by treating XIX with N-butyllithium, lithium diisopropylamine or sodium hydride, at a temperature of about xe2x88x9278xc2x0 C., in the presence of a polar aprotic solvent, such as tetrahydrofuran. The anionic intermediate so formed is further reacted with (a) alkylhalide or benzylhalide, at a temperature between about xe2x88x9278xc2x0 C. to room temperature, preferably xe2x88x9278xc2x0 C., when R3 is alkyl or benzyl; (b) an aldehyde or ketone, at a temperature between about xe2x88x9278xc2x0 C. to room temperature, preferably xe2x88x9278xc2x0 C., when R3 is alkoxy; and (c) zinc chloride, at a temperature between about xe2x88x9278xc2x0 C. to room temperature, preferably xe2x88x9278xc2x0 C., and the corresponding organozinc intermediate so formed is then reacted with aryliodide or vinyl iodide in the presence of a catalytic quantity of palladium. The resulting reaction mixture is stirred at a temperature between about 50xc2x0 C. to about 80xc2x0 C., preferably about 70xc2x0 C., for a time period between about 1 hour to about 3 hours, preferably about 1 hour. Alternatively, the anion so formed is reacted with molecular oxygen to form the corresponding 4-chloro-6-hydroxypyrrolo[2,3-d]pyrimidine compound of formula XVI, wherein R3 is hydroxy.
In reaction 1 of Scheme 3, the 4-chloropyrrolo[2,3-d]pyrimidine compound of formula XXI is converted to the corresponding compound of formula XXII, according to the procedure described above in reaction 3 of Scheme 2.
In reaction 2 of Scheme 3, the compound of formula XXII is converted to the corresponding compound of formula XVI, according to the procedures described above in reactions 1 and 2 of Scheme 3.
In reaction 1 of Scheme 4, the 4-chloropyrrolo[2,3-d]pyrimidine compound of formula XX is converted to the corresponding 4-aminopyrrolo[2,3-d]pyrimidine compound of formula XXIV, according to the procedure described above in reaction 2 of Scheme 1.
In reaction 2 of Scheme 4, the 4-amino-5-halopyrrolo[2,3-d]pyrimidine compound of formula XXIV, wherein R is benzenesulfonate and Z is bromine or iodine, is converted to the corresponding compound of formula XXIII by reacting XXIV with (a) arylboronic acid, when R2 is aryl, in an aprotic solvent, such tetrahydrofuran or dioxane, in the presence of a catalytic quantity. of palladium (0) at a temperature between about 50xc2x0 C. to about 100xc2x0 C., preferably about 70xc2x0 C., for a time period between about 2 hours to about 48 hours, preferably about 12 hours; (b) alkynes, when R2 is alkynyl, in the presence of acatalytic quantity of copper (I) iodide and palladium (0), and a polar solvent, such as dimethylformamide, at room temperature, for a time period between about 1 hour to about 5 hours, preferably about 3 hours; and (c) alkenes or styrenes, when R2 is vinyl or styrenyl, in the presence of a catalytic quantity of palladiumn in dimethylformamide, dioxane or tetrahydrofuran, at a temperature between about 80xc2x0 C. to about 100xc2x0 C., preferably about 100xc2x0 C., for a time period between about 2 hours to about 48 hours, preferably about 48 hours.
In reaction 3 of Scheme 4, the compound of formula XXIII is converted to the corresponding compound of formula XV, according to the procedure described above in reaction 3 of Scheme 2.
The compounds of the present invention that are basic in nature are capable of forming a wide variety of different salts with various. inorganic and organic acids. Although such salts must be pharmaceutically acceptable for administration to animals, it is often desirable in practice to initially isolate the compound of the present invention from the reaction mixture as a pharmaceutically unacceptable salt and then simply convert the latter back to the free base compound by treatment with an alkaline reagent and subsequently convert the latter free base to a pharmaceutically acceptable acid addition salt. The acid addition salts of the base compounds of this invention are readily prepared by treating the base compound with a substantially equivalent amount of the chosen mineral or organic acid in an aqueous solvent medium or in a suitable organic solvent, such as methanol or ethanol. Upon careful evaporation of the solvent, the desired solid salt is readily obtained. The desired acid salt can also be precipitated from a solution of the free base in an organic solvent by adding to the solution an appropriate mineral or organic acid.
Those compounds of the present invention that are acidic in nature, are capable of forming base salts with various pharmacologically acceptable cations. Examples of such salts include the alkali metal or alkaline-earth metal salts and particularly, the sodium and potassium salts. These salts are all prepared by conventional techniques. The chemical bases which are used as reagents to prepare the pharmaceutically acceptable base salts of this invention are those which form non-toxic base salts with the acidic compounds of the present invention. Such non-toxic base salts include those derived from such pharmacologically acceptable cations as sodium, potassium calcium and magnesium, etc. These salts can easily be prepared by treating the corresponding acidic compounds with an aqueous solution containing the desired pharmacologically acceptable cations, and then evaporating the resulting solution to dryness, preferably under reduced pressure. Alternatively, they may also be prepared by mixing lower alkanolic solutions of the acidic compounds and the desired alkali metal alkoxide together, and then evaporating the resulting solution to dryness in the same manner as before. In either case, stoichiometric quantities of reagents are preferably employed in order to ensure completeness of reaction and maximum yields of the desired final product.
The compositions of the present invention may be formulated in a conventional manner using one or more pharmaceutically acceptable carriers. Thus, the active compounds of the invention may be formulated for oral, buccal, intranasal, parenteral (eg, intravenous, intramuscular or subcutaneous) or rectal administration or in a form suitable for administration by inhalation or insufflation. The active compounds of the invention may also be formulated for sustained delivery.
For oral administration, the pharmaceutical compositions may take the form of, for example, tablets or capsules prepared by conventional means with pharmaceutically acceptable excipients such as binding agents (e.g., pregelatinized maize starch, polyvinylpyrrolidone or hydroxypropyl methylcellulose); fillers (e.g., lactose, microcrystalline cellulose or calcium phosphate); lubricants (e.g., magnesium stearate, talc or silica); disintegrants (e.g., potato starch or sodium starch glycolate); or wetting agents (e.g., sodium lauryl sulphate). The tablets may be coated by methods well known in the art. Liquid preparations for oral administration may take the form of, for example, solutions, syrups or suspensions, or they may be presented as a dry product for constitution with water or other suitable vehicle before use. Such liquid preparations may be prepared by conventional means with pharmaceutically acceptable additives such as suspending agents (e.g., sorbitol syrup, methyl cellulose or hydrogenated edible fats); emulsifying agents (e.g., lecithin or acacia); non-aqueous vehicles (e.g., almond oil, oily esters or ethyl alcohol); and preservatives (e.g., methyl or propyl p-hydroxybenzoates or sorbic acid).
For buccal administration, the composition may take the form of tablets or lozenges formulated in conventional manner.
The active compounds of the invention may be formulated for parenteral administration by injection, including using conventional catheterization techniques or infusion. Formulations for injection may be presented in unit dosage form, e.g., in ampules or in multi-dose containers, with an added preservative. The compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulating agents such as suspending, stabilizing and/or dispersing agents. Alternatively, the active ingredient may be in powder form for reconstitution with a suitable vehicle, e.g., sterile pyrogen-free water, before use.
The active compounds of the invention may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter or other glycerides.
For intranasal administration or administration by inhalation, the active compounds of the invention are conveniently delivered in the form of a solution or suspension from a pump spray container that is squeezed or pumped by the patient or as an aerosol spray presentation from a pressurized container or a nebulizer, with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. The pressurized container or nebulizer may contain a solution or suspension of the active compound. Capsules and cartridges (made, for example, from gelatin) for use in an inhaler or insufflator may be formulated containing a powder mix of a compound of the invention and a suitable powder base such as lactose or starch.
A proposed dose of the active compounds of the invention for oral, parenteral or buccal administration to the average adult human for the treatment of the conditions referred to above (e.g., asthma) is 0.1 to 1000 mg of the active ingredient per unit dose which could be administered, for example, 1 to 4 times per day.
Aerosol formulations for treatment of the conditions referred to above (es, rheumatoid arthritis) in the average adult human are preferably arranged so that each metered dose or xe2x80x9cpuffxe2x80x9d of aerosol contains 20 xcexcg to 1000 xcexcg of the compound of the invention. The overall daily dose with an aerosol will be within the range 0.1 mg to 1000 mg. Administration may be several times daily, for example 2, 3, 4 or 8 times, giving for example, 1, 2 or 3 doses each time.
A compound of formula (I) administered in a pharmaceutically acceptable form either alone or in combination with one or more additional agents which modulate a mammlian immune system or with antiinflammatory agents, agents which may include but are not limited to cyclosporin A (e.g. Sandimmunec(copyright) or Neoral(copyright), rapamycin, FK-506 (tacrolimus), leflunomide, deoxyspergualin, mycophenolate (e.g. Cellcept(copyright)), azathioprine (e.g. Imuran(copyright)), daclizumab (e.g. Zenapax(copyright)), OKT3 (e.g. Orthocolone(copyright)), AtGam, aspirin, acctaminophen, ibuprofen, naproxen, piroxicam, and antiinflmmatory steroids (e.g. prednisolone or dexamethasone); and such agents may be administered as part of the same or separate dosage forms, via the same or different routes of administration, and on the same or different administration schedules according to standard pharmaceutical practice.
FK506 (Tacrolimus) is given orally at 0.10-0.15 mg/kg body weight, every 12 hours, within first 48 hours postoperative. Does is monitored by serum Tacrolimus trough levels. Cyclosporin A (Sandimmune oral or intravenous formulation, or Neoral(copyright), oral solution or capsules) is given orally at 5 mg/kg body weight, every 12 hours within 48 hours postoperative. Dose is monitored by. blood Cyclosporin A trough levels.
The active agents can be formulated for sustained delivery according to methods well known to those of ordinary skill in the art. Examples of such formulations can be found in U.S. Pat. Nos. 3,538,214, 4,060,598, 4,173,626, 3,119,742, and 3,492,397.
The ability of the compounds of formula I or their pharmaceutically acceptable salts to inhibit Janus Kinase 3 and, consequently, demonstrate their effectiveness for treating disorders or conditions characterized by Janus Kinase 3 is shown by the following in vitro assay tests.
JAK3 (JH1:GST) Enzymatic Assay
The JAK3 kinase assay utilizes a protein expressed in baculovirus-infected SF9 cells (a fusion protein of GST and the catalytic domain of human JAK3) purified by affinity chromatography on glutathione-Sepaharose. The substrate for the reaction is poly-Glutamic acid-Tyrosine (PGT (4:1), Sigma catalog #P0275), coated onto Nunc Maxi Sorp plates at 100 xcexc/ml overnight at 37xc2x0 C. The morning after coating, the plates are washed three times and JAK3 is added to the wells containing 100 xcexcl of kinase buffer (50 mM HEPES, pH 7.3, 125 mM NaCl, 24 mM MgCl2)+0.2 uM ATP+1 mM Na orthovanadate.) The reaction proceeds for 30 minutes at room temperature and the plates is washed three more times. The level of phosphorylated tyrosine in a given well is quantitated by standard ELISA assay utilizing an anti-phosphotyrosine antibody (ICN PY20, cat. #69-151-1).
DND 39/IL-4 Cellular Assay for JAK3 kinase Inhibitors
The DND 39/IL-4 assay is designed to find inhibitors of JAK3 kinase activity which would be prime candidates for immunosupressive and/or allergy. The assay uses a B-cell line called DND39 which has had the luciferase gene driven by the germ line IgE promoter stably integrated into one of the chromosomes. When these cells are stimulated with IL-4, the kinase JAK3, which is associated with the IL-4 receptor, phosphorylates the signal transducer STAT6. STAT6 then blinds to the germline IgE promoter and starts transcription of the luciferase gene. Luciferase is measured in a lysate of these cells using the Promega luciferase assay reagent system.
Note: DND39 cells are grown in RPMI 1640 supplemented with 10% heat inactivated FCS, 2 mM L-Glutamine, and 100 units/ml Pen./Strep. The cells are maintained from 1xc3x97105 to 1xc3x97106 cells/ml. Split to 1xc3x97105 on Friday, cells will be at about 1xc3x97106 on Monday. Then split 1:2 during the week keeping 200 ml in a flask as needed.
3xc3x97105 DND39 cells are plated in 100 xcexcl of RPMI 1640 supplemented with 1% heat inactivated FCS, 2 mM L-glutamine, and 100 units/ml Pen/Step in a 96 well Vee bottom plate (Nunc). Compounds are diluted serially 1:2 in DMSO starting at 4 mM to 1.9 xcexcM. In a 96 well polypropylene plate, changing tips after each dilution. Then 5 xcexcl of each dilution are added to 500 xcexcl of RPMI/1% serum in a 96 tube rack. 125 xcexcL of the compound dilutions are added to the cells and incubated at 37xc2x0 C., 5% CO2 for one hour. After one hour, 25 xcexcl of 25 ng/ml IL-4 is added to the cells and mixed. Final concentration of IL-4 is 2.5 ng/ml and final concentration of compound is from 20 xcexcM to 156 nM. The cells are then incubated overnight 16-18 hours. The plate is then centrifuged at 2500-3000 RPM in a table top centrifuge for 5 minutes. The culture supernatant is carefully removed by aspiration with an 8 well maifold. 100 xcexcl of PBS with calcium and magnesium is added to the pelletted cells. The cells are resuspended in the PBS and transferred to a Packard white OptiPlate. 100 xcexcl of Packard""s LucLite reagent is added to the wells of the OptiPlate.
The following Examples illustrate the preparation of the compounds of the present invention but it is not limited to the details thereof. Melting points are uncorrected. NMR data are reported in parts per million (xcex4) and are referenced to the deuterium lock signal from the sample solvent (deuteriochloroform unless otherwise specified). Commercial reagents were utilized without further purification. THF refers to tetrahydrofuran. DMF refers to N,N-dimethylformamide. Low Resolution Mass Spectra (LRMS) were recorded on either a Hewlett Packard 5989(copyright), utilizing chemical ionization (ammonium), or a Fisons (or Micro Mass) Atmospheric Pressure Chemical Ionization (APCI) platform which uses a 50/50 mixture of acetonitrile/water with 0.1% formic acid as the ionizing agent. Room or ambient temperature refers to 20-25xc2x0 C.